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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 505 records · Page 28
All sourcesNASA NTRSDOE OSTI
NASA NTRS2001

Focal adhesion kinase is involved in mechanosensing during fibroblast migration

Focal adhesion kinase (FAK) is a non-receptor protein tyrosine kinase localized at focal adhesions and is believed to mediate adhesion-stimulated effects. Although ablation of FAK impairs cell movement, it is not clear whether FAK might be involved in the guidance of cell migration, a role consistent with its putative regulatory function. We have transfected FAK-null fibroblasts with FAK gene under the control of the tetracycline repression system. Cells were cultured on flexible polyacrylamide substrates for the detection of traction forces and the application of mechanical stimulation. Compared with control cells expressing wild-type FAK, FAK-null cells showed a decrease in migration speed and directional persistence. In addition, whereas FAK-expressing cells responded to exerted forces by reorienting their movements and forming prominent focal adhesions, FAK-null cells failed to show such responses. Furthermore, FAK-null cells showed impaired responses to decreases in substrate flexibility, which causes control cells to generate weaker traction forces and migrate away from soft substrates. Cells expressing Y397F FAK, which cannot be phosphorylated at a key tyrosine site, showed similar defects in migration pattern and force-induced reorientation as did FAK-null cells. However, other aspects of F397-FAK cells, including the responses to substrate flexibility and the amplification of focal adhesions upon mechanical stimulation, were similar to that of control cells. Our results suggest that FAK plays an important role in the response of migrating cells to mechanical input. In addition, phosphorylation at Tyr-397 is required for some, but not all, of the functions of FAK in cell migration.

NASA Discipline Cell Biology↗
NASA NTRS2003

Multilineage differentiation of rhesus monkey embryonic stem cells in three-dimensional culture systems

In the course of normal embryogenesis, embryonic stem (ES) cells differentiate along different lineages in the context of complex three-dimensional (3D) tissue structures. In order to study this phenomenon in vitro under controlled conditions, 3D culture systems are necessary. Here, we studied in vitro differentiation of rhesus monkey ES cells in 3D collagen matrixes (collagen gels and porous collagen sponges). Differentiation of ES cells in these 3D systems was different from that in monolayers. ES cells differentiated in collagen matrixes into neural, epithelial, and endothelial lineages. The abilities of ES cells to form various structures in two chemically similar but topologically different matrixes were different. In particular, in collagen gels ES cells formed gland-like circular structures, whereas in collagen sponges ES cells were scattered through the matrix or formed aggregates. Soluble factors produced by feeder cells or added to the culture medium facilitated ES cell differentiation into particular lineages. Coculture with fibroblasts in collagen gel facilitated ES cell differentiation into cells of a neural lineage expressing nestin, neural cell adhesion molecule, and class III beta-tubulin. In collagen sponges, keratinocytes facilitated ES cell differentiation into cells of an endothelial lineage expressing factor VIII. Exogenous granulocyte-macrophage colony-stimulating factor further enhanced endothelial differentiation. Thus, both soluble factors and the type of extracellular matrix seem to be critical in directing differentiation of ES cells and the formation of tissue-like structures. Three-dimensional culture systems are a valuable tool for studying the mechanisms of these phenomena.

Pluripotent Stem Cells/cytology/physiology↗
NASA NTRS1999

Induction of three-dimensional assembly of human liver cells by simulated microgravity

The establishment of long-term cultures of functional primary human liver cells (PHLC) is formidable. Developed at NASA, the Rotary Cell Culture System (RCCS) allows the creation of the unique microgravity environment of low shear force, high-mass transfer, and 3-dimensional cell culture of dissimilar cell types. The aim of our study was to establish long-term hepatocyte cultures in simulated microgravity. PHLC were harvested from human livers by collagenase perfusion and were cultured in RCCS. PHLC aggregates were readily formed and increased up to 1 cm long. The expansion of PHLC in bioreactors was further evaluated with microcarriers and biodegradable scaffolds. While microcarriers were not conducive to formation of spheroids, PHLC cultured with biodegradable scaffolds formed aggregates up to 3 cm long. Analyses of PHLC spheroids revealed tissue-like structures composed of hepatocytes, biliary epithelial cells, and/or progenitor liver cells that were arranged as bile duct-like structures along nascent vascular sprouts. Electron microscopy revealed groups of cohesive hepatocytes surrounded by complex stromal structures and reticulin fibers, bile canaliculi with multiple microvilli, and tight cellular junctions. Albumin mRNA was expressed throughout the 60-d culture. A simulated microgravity environment is conducive to maintaining long-term cultures of functional hepatocytes. This model system will assist in developing improved protocols for autologous hepatocyte transplantation, gene therapy, and liver assist devices, and facilitate studies of liver regeneration and cell-to-cell interactions that occur in vivo.

NASA Center JSC↗
NASA NTRS1998

Cell contact as an independent factor modulating cardiac myocyte hypertrophy and survival in long-term primary culture

Cardiac myocytes maintained in cell culture develop hypertrophy both in response to mechanical loading as well as to receptor-mediated signaling mechanisms. However, it has been shown that the hypertrophic response to these stimuli may be modulated through effects of intercellular contact achieved by maintaining cells at different plating densities. In this study, we show that the myocyte plating density affects not only the hypertrophic response and features of the differentiated phenotype of isolated adult myocytes, but also plays a significant role influencing myocyte survival in vitro. The native rod-shaped phenotype of freshly isolated adult myocytes persists in an environment which minimizes myocyte attachment and spreading on the substratum. However, these conditions are not optimal for long-term maintenance of cultured adult cardiac myocytes. Conditions which promote myocyte attachment and spreading on the substratum, on the other hand, also promote the re-establishment of new intercellular contacts between myocytes. These contacts appear to play a significant role in the development of spontaneous activity, which enhances the redevelopment of highly differentiated contractile, junctional, and sarcoplasmic reticulum structures in the cultured adult cardiomyocyte. Although it has previously been shown that adult cardiac myocytes are typically quiescent in culture, the addition of beta-adrenergic agonists stimulates beating and myocyte hypertrophy, and thereby serves to increase the level of intercellular contact as well. However, in densely-plated cultures with intrinsically high levels of intercellular contact, spontaneous contractile activity develops without the addition of beta-adrenergic agonists. In this study, we compare the function, morphology, and natural history of adult feline cardiomyocytes which have been maintained in cultures with different levels of intercellular contact, with and without the addition of beta-adrenergic agonists. Intercellular contact, communication, and transmission of contractile forces between myocytes appears to play a primary role in remodeling the 2-dimensional cell layer into a parallel alignment of elongated myocytes with highly developed intercalated disk-like junctions. This highly differentiated state is very stable, and cultures which achieve this state exhibit significantly greater longevity than more sparsely plated myocytes. These myocytes typically continue beating, and survive from 6 to more than 12 weeks in culture. When this level of contact and differentiation are not achieved, even among beta-adrenergic stimulated myocytes, contractile activity is not sustained, myofibrils atrophy, there is little or no development of junctional complexes, and the period of myocyte viability is typically no more than 5 weeks in vitro.

Non-NASA Center↗
NASA NTRS1992

Friend leukemia virus transformed cells exposed to microgravity in the presence of DMSO (7-IML-1)

The purpose of this experiment is to study the adaptation of living cells to microgravity. The in vitro transformation of Friend cells by Dimethylsufoxide (DMSO) is a good model for the study of cell differentiation and protein biosynthesis. Cultures of cells will be prepared shortly before launch. Once in space, transformation will be induced by injection of DMSO. One set of cultures will be chemically fixed with glutaraldehyde for electron microscope investigations; another set will be preserved for determining the amount of hemogloben produced and the extent of cell proliferation.

Cogoli, Augusto↗
NASA NTRS1984

Cells, embryos and development in space

Work continues to focus on the demonstrable totipotency of cultured somatic cells of various higher plants and has examined the conditions which regulate this propensity to be controllably released. This was done with special reference to cells obtained from cultured explants of daylily and carrot. For purposes of identifying the variables in question, work was carried out almost exclusively in liquid media. The events that intervene between the aseptic isolation of tissue explants, the culture of small derived units and free cells and the propagation in large numbers of adventive or somatic embryos to plantlets were traced and certain definitive stages at which control is exercised were identified. In daylily, morphologically competent units are now propagated with a high degree of precision in rotated liquid cultures in bulk, and under the conditions of continuous neutralized gravity, the development progresses so that embryo-plantlets are obtained.

Krikorian, A. D.↗
NASA NTRS1985

Kidney cell electrophoresis

Tasks were undertaken in support of two objectives. They are: (1) to carry out electrophoresis experiments on cells in microgravity; and (2) assess the feasibility of using purified kidney cells from embryonic kidney cultures as a source of important cell products. Investigations were carried out in the following areas: (1) ground based electrophoresis technology; (2) cell culture technology; (3) electrophoresis of cells; (4) urokinase assay research; (5) zero-g electrophoresis; and (6) flow cytometry.

Todd, P. W.↗
NASA NTRS2004

The Biotechnology Facility for International Space Station

The primary mission of the Cellular Biotechnology Program is to advance microgravity as a tool in basic and applied cell biology. The microgravity environment can be used to study fundamental principles of cell biology and to achieve specific applications such as tissue engineering. The Biotechnology Facility (BTF) will provide a state-of-the-art facility to perform cellular biotechnology research onboard the International Space Station (ISS). The BTF will support continuous operation, which will allow performance of long-duration experiments and will significantly increase the on-orbit science throughput.

NASA Center JSC↗
NASA NTRS1998

Taxane recovery from cells of Taxus in micro- and hypergravity

Cell suspension cultures of Taxus cuspidata produce taxanes that are released from the outer surface of cells into the culture medium as free and bound alkaloids. Paclitaxel (Taxol (TM)), is an anti-cancer drug in short supply. It has a taxane ring derived from baccatin III and a C-13 phenylisoserine side-chain. This drug is produced over a wide range of gravitational forces. Monoclonal and polyclonal antibodies to paclitaxel, baccatin III, and the C-13 phenylisoserine side chain were combined in multiple-labeling studies to localize taxanes and paclitaxel on cell surfaces or on particles released into the culture medium. Bioreactor vessel design altered the composition of taxanes recovered from cells in simulated microgravity. At 10(-2) and 2x10(-4)g, taxane recovery was reduced but biomass growth and percent paclitaxel was significantly increased. At 1 to 24g, growth was reduced with a significant recovery of total taxanes with low percent paclitaxel. Bound paclitaxel was also localized in endonuclease-rich fragmenting nuclei of individual apoptotic cells. A model is presented comprising TCH (touch) genes encoding enzymes that modify taxane-bearing xylan residues in cell walls, the calcium-sensing of gravitational forces by the cytoplasm, and the predisposition of nuclei to apoptosis. This integrates the adaptive physiological and biochemical responses of drug-producing genomes with gravitational forces.

Plants, Medicinal/chemistry/cytology↗
NASA NTRS2000

Particle irradiation induces FGF2 expression in normal human lens cells

Particle Irradiation Induces FGF2 Expression in Normal Human Lens Cells. Particle radiations, including both proton and helium-ion beams, have been used to successfully treat choroidal melanoma, but with the complication of radiation-induced cataract. We have investigated a role for radiation-induced changes in the expression of basic fibroblast growth factor (FGF2) gene expression as part of the mechanism(s) underlying lens cell injury associated with cataract. Normal human lens epithelial (HLE) cells were cultured in vitro on extracellular matrix (ECM) originated from bovine corneal endothelial cells. This study reports evidence for rapid but transient induction of FGF2 transcripts, an increase of between 5- and 8-fold, within 0.5 h after exposure to particle radiation, followed by another wave of increased transcription at 2-3 h postirradiation. Immunofluorescence results confirm the enhanced levels of FGF2 protein rapidly after exposure to protons or helium ions, followed by another wave of increased activity unique to helium at 6 h postirradiation. This second wave of increased immunoreactivity was not observed in the proton-irradiated samples. Total FGF2 protein analysis after helium-ion exposures shows induced expression of three FGF2 isoforms, with an increase of up to 2-fold in the 18-kDa low-molecular-weight species. Studies of the effects of protons on individual FGF2 protein isoforms are in progress. Several mechanisms involving a role for FGF2 in radiation-induced cataract are discussed.

NASA Discipline Radiation Health↗
NASA NTRS1999

Effects of space flight and mixing on bacterial growth in low volume cultures

Previous investigations have shown that liquid suspension bacterial cultures grow to higher cell concentrations in spaceflight than on Earth. None of these studies included ground-control experiments designed to evaluate the fluid effects potentially responsible for the reported increases. Therefore, the emphasis of this research was to both confirm differences in final cell concentration between 1g and microgravity cultures, and to examine the effects of mixing as a partial explanation for this difference. Flight experiments were performed in the Fluid Processing Apparatus (FPA), aboard Space Shuttle Missions STS-63 and STS-69, with simultaneous 1g static and agitated controls. Additional static 1g, agitated, and clino-rotated controls were performed in 9-ml culture tubes. This research revealed that both E. coli and B. subtilis samples cultured in space flight grew to higher final cell densities (120-345% increase) than simultaneous static 1g controls. The final cell concentration of E. coli cells cultured under agitation was 43% higher than in static 1g cultures and was 102% higher with clino-rotation. However, for B. subtilis cultures grown while being agitated on a shaker or clino-rotated, the final cell concentrations were nearly identical to those of the simultaneous static 1g controls. Therefore, these data suggest that the unique fluid quiescence in the microgravity environment (lack of sedimentation, creating unique transfer of nutrients and waste products), was responsible for the enhanced bacterial proliferation reported in this and other studies.

STS-63 Shuttle Project↗
NASA NTRS2006

An Optimization of Pulsed ElectroMagnetic Fields Study

To date, in our research we have focused on the use of normal human neuronal progenitor (NHNP) cells because of their importance in human nervous system regeneration, development and maintenance, but we have developed 2-D and 3-D bioreactors that can accommodate any cell line. In this Project, we will include the use of tissues important for physiological regeneration: Human osteoblasts or chondrocytes, and vascular cells. Our initial results with the NHNP cells were quite startling using extremely low-level electromagnetic fields (5 microtesla at 10Hz; 6mA). The low-amplitude, rapidly time-varying electromagnetic fields exert a very potent effect on the proliferation, morphology, and gene expression of the cells in culture, both in standard 2-dimensional culture plates as well as cells organized into 3-dimensional tissue-like assemblies (TLAs) in a 3D bioreactor. We have replicated our preliminary results many, many times, have analyzed the gene expression using gene arrays (followed by Luminex analysis for protein production), and have monitored cell proliferation, orientation, morphology, and glucose metabolism, and we are confident that we have a stable and reliable model to study the control of high-level cellular processes by application of low-amplitude, time varying electromagnetic fields (TVEMF) (1, 2). In additional studies at the University of Michigan, we have been able to generate functional in vitro engineered mammalian skeletal muscle, and have employed nerve-muscle co-culture techniques to promote axonal sprouting. We believe that nearly all tissues, in particular, neural, are susceptible to the influences of low-level TVEMF.

Goodwin, Thomas J.↗
NASA NTRS2005

Tissue Engineering Using Transfected Growth-Factor Genes

A method of growing bioengineered tissues includes, as a major component, the use of mammalian cells that have been transfected with genes for secretion of regulator and growth-factor substances. In a typical application, one either seeds the cells onto an artificial matrix made of a synthetic or natural biocompatible material, or else one cultures the cells until they secrete a desired amount of an extracellular matrix. If such a bioengineered tissue construct is to be used for surgical replacement of injured tissue, then the cells should preferably be the patient s own cells or, if not, at least cells matched to the patient s cells according to a human-leucocyteantigen (HLA) test. The bioengineered tissue construct is typically implanted in the patient's injured natural tissue, wherein the growth-factor genes enhance metabolic functions that promote the in vitro development of functional tissue constructs and their integration with native tissues. If the matrix is biodegradable, then one of the results of metabolism could be absorption of the matrix and replacement of the matrix with tissue formed at least partly by the transfected cells. The method was developed for articular chondrocytes but can (at least in principle) be extended to a variety of cell types and biocompatible matrix materials, including ones that have been exploited in prior tissue-engineering methods. Examples of cell types include chondrocytes, hepatocytes, islet cells, nerve cells, muscle cells, other organ cells, bone- and cartilage-forming cells, epithelial and endothelial cells, connective- tissue stem cells, mesodermal stem cells, and cells of the liver and the pancreas. Cells can be obtained from cell-line cultures, biopsies, and tissue banks. Genes, molecules, or nucleic acids that secrete factors that influence the growth of cells, the production of extracellular matrix material, and other cell functions can be inserted in cells by any of a variety of standard transfection techniques.

Madry, Henning↗
NASA NTRS1985

Microbial colonization and growth on metal sulfides and other mineral surfaces

To determine whether a bacterial film forms on sulfur minerals in situ, various sulfur containing and other minerals were incubated in Penitencia Creek. The rate of cell growth and attachment within the surface microenvironment of mineral surfaces was also determined. To determine whether surfaces enriched with soluble sulfur substrates (cysteine, glutathione, thioglycolate, sulfite, and thiosulfate) increased the rate of growth or attachment of natural communities, membrane enrichments were incubated. These rates were determined as described by Caldwell et al. (1981, 1983). The growth of Pseudomonas fluorescens, a heterotrophic sulfur oxidizer, was studied in batch cell suspensions and in continuous culture. In batch culture the cells were oxygen limited (growth rate 0.33 per hour under oxygen limitations and 0.52 per hour when vigorously aerated). Growth within the film was glucose limited. Several behavioral phenomena were observed for cells growing within the hydrodynamic boundary layer. Despite a flow of 10 cm per second in the environment, the bacteria were able to move freely in both directions within the hydrodynamic boundary layer.

Caldwell, D.↗
NASA NTRS1993

Characterization of Spirulina biomass for CELSS diet potential

Cyanobacteria, Spirulina maxima as a biogenerative photosynthetic and an edible alga for the space craft crew in a CELSS, was evaluated in an effort to increase the growth rate, biomass, yield, and chemical analysis in continuous cultures. The cell characteristics were determined for cultures maintained at steady state with respect to the substrate concentration. The productivity increased in experiments exposed to low light (30 uE m(exp -2)s(exp -1). Oxygen evolved and protein production were higher in cultures exposed to low light intensity. There was a relationship between nitrate concentration and the yield of the culture. Increasing the concentration of nitrate in the growth medium up to 20 mM was enough to produce a culture having the same chemical composition as that of complete medium. High light was inhibiting the yield of the culture. Increasing the concentration of phosphate beyond 1 mM did not improve the yield of the culture. Increasing the concentration of sodium chloride in the growth medium did not affect the growth of the alga up to 0.1 M but beyond that the culture started to be stressed. The response to stress appeared in high production of total carbohydrate on the expense of protein production. The oxygen production was also higher in cultures stressed with sodium chloride.

Tadros, Mahasin G.↗
NASA NTRS1994

Experiment K-7-23: Effect of Spaceflight on Level and Function of Immune Cells: Proliferation and Cytokines - Part 2

Lymphocytes from the superficial inguinal lymph nodes of rats flown on the Cosmos 2044 space mission were tested for proliferation in response to polyclonal activators. Cells were cultured with T or B cell mitogens, phorbol ester and calcium ionophore, or T cell mitogen and the lymphokines interleukin-1 (IL-1) or interleukin-2 (IL-2), and assayed for DNA synthesis by (3)H-thymidine incorporation. Lymphocytes also were incubated with concanavalin A (Con A), a T cell mitogen, and tested for IL-2 production. Mitogen-stimulated proliferation of lymphocytes from rats exposed to microgravity was not significantly different from synchronous or vivarium controls. Responses to Con A and IL-2, and Con A and IL-1 likewise were unaffected by space flight. Lymphocytes from all of these groups responded well to phorbol ester and calcium ionophore stimulation. Furthermore, lymph node cells (LNC) from control rats and rats flown on Cosmos 2044 produced similar amounts of IL-2. The results obtained using hindlimb suspended rats were notably different from those of flight and control animals. LNC from suspended rats generally had greater proliferative responses to T cell mitogens than did lymphocytes from other groups. Responsiveness to a B cell mitogen was not enhanced. Con A-stimulated LNC from hindlimb suspended rats also produced more IL-2 than did lymphocytes from the other groups. This difference was statistically significant at both IL-2 induction times tested.

Nash, P. V.↗
NASA NTRS1989

Improved method for HPLC analysis of polyamines, agmatine and aromatic monoamines in plant tissue

The high performance liquid chromatographic (HPLC) method of Flores and Galston (1982 Plant Physiol 69: 701) for the separation and quantitation of benzoylated polyamines in plant tissues has been widely adopted by other workers. However, due to previously unrecognized problems associated with the derivatization of agmatine, this important intermediate in plant polyamine metabolism cannot be quantitated using this method. Also, two polyamines, putrescine and diaminopropane, also are not well resolved using this method. A simple modification of the original HPLC procedure greatly improves the separation and quantitation of these amines, and further allows the simulation analysis of phenethylamine and tyramine, which are major monoamine constituents of tobacco and other plant tissues. We have used this modified HPLC method to characterize amine titers in suspension cultured carrot (Daucas carota L.) cells and tobacco (Nicotiana tabacum L.) leaf tissues.

Non-NASA Center↗
NASA NTRS2014

High-Density Spot Seeding for Tissue Model Formation

A method for making a tissue includes seeding cells at a selected concentration on a support to form a cell spot, incubating the cells to allow the cells to partially attach, rinsing the cells to remove any unattached cells, adding culture medium to enable the cells to proliferate at a periphery of the cell spot and to differentiate toward a center of the cell spot, and further incubating the cells to form the tissue. The cells may be C2C12 cells or other subclones of the C2 cell line, H9c2(2-1) cells, L6 cells, L8 cells, QM7 cells, Sol8 cells, G-7 cells, G-8 cells, other myoblast cells, cells from other tissues, or stem cells. The selected concentration is in a range from about 1 x 10(exp 5) cells/ml to about 1 x 10(exp 6) cells/ml. The tissue formed may be a skeletal muscle tissue, a cardiac muscle tissue, nerve tissue, or a bone tissue.

Marquette, Michele L.↗